The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details respecting the practice, are incorporated herein by reference, and for convenience, are referenced by author and date in the following text and respectively grouped in the appended List of References.
The genetics of cancer is complicated, involving the function of three loosely defined classes of genes: (1) dominant, positive regulators of the transformed state (oncogenes); (2) recessive, negative regulators of the transformed state (tumor suppressor genes); and (3) genes that modify risk without playing a direct role in the biology of transformed cells (risk modifiers).
Specific germline alleles of certain oncogenes and tumor suppressor genes are causally associated with predisposition to cancer. This set of genes is referred to as tumor predisposition genes. Some of the tumor predisposition genes which have been cloned and characterized influence susceptibility to: 1) Retinoblastoma (RB1); 2) Wilms' tumor (WT1); 3) Li-Fraumeni (TP53); 4) Familial adenomatous polyposis (APC); 5) Neurofibromatosis type 1 (NF1); 6) Neurofibromatosis type 2 (NF2); 7) von Hippel-Lindau syndrome (VHL); 8) Multiple endocrine neoplasia type 2A (MEN2A); 9) Melanoma (CDKN2 and CDK4); 10) Breast and ovarian cancer (BRCA1 and BRCA2); 11) Cowden disease (MMAC1); 12) Multiple endocrine neoplasia (MEN1); 13) Nevoid basal cell carcinoma syndrome (PTC); 14) Tuberous sclerosis 2 (TSC2); 15) Xeroderma pigtpentosum (genes involved in nucleotide excision repair); 16) Hereditary nonpolyposis colorectal cancer (genes involved in mismatch repair).
Specific germline alleles of certain risk modifier genes arc also associated with predisposition to cancer, but the increased risk is sometimes only clearly expressed when it is combined with certain environmental, dietary, or other factors. Alcohol dehydrogenase CADH) oxidizes ethanol to acetaldehyde, a chemical which is both mutagenic and carcinogenic in lab animals. The enzyme encoded by the ADH3.sup.1 allele oxidizes ethanol relatively quickly, whereas the enzyme encoded by the ADH3.sup.2 allele oxidizes ethanol more slowly. ADH3.sup.1 homozygotes presumably have a high capacity for synthesis of acetaldehyde; those who also drink heavily are at increased risk for oral cavity, esophageal, and (in women) breast cancer relative to ADH3.sup.2 homozygotes who drink equally heavily (Harty et al., 1997; Hori et al., 1997; Shields, 1997). The acetyltransferases encoded by N-acetyltransferase 1 (NAT1) and N-acetyltransferase 2 (NAT2) catalyze the acetylation of numerous xenobiotics including the aromatic amine carcinogens derived from smoking tobacco products. Individuals who are homozygous for slow acetylating forms of NAT2 who are also heavy smokers are at greater risk for lung, bladder, and (in females) breast cancer than individuals who smoke equally heavily but are homozygous for fast acetylating forms of NAT2 (Shields, 1997; 13ouchardy et al., 1998).
The risk of hormone related cancers such as breast and prostate cancer may be modulated by allelic variants in enzymes that play a role in estrogen or androgen metabolism, or variants in proteins that mediate the biological effects of estrogens or androgens. A polymoiphic CAG repeat in the human androgen receptor gene encodes a polymorphic polyglutamine tract near the amino-terminus of the protein. The length of the polyglutamine tract is inversely correlated with the transcriptional activation activity of the androgen receptor and thus one aspect of the biological response to androgens. Men whose androgen receptor contains a relatively short polyglutamine tract are at higher risk for prostate cancer, especially high stage/high histologic grade prostate cancer, than men whose androgen receptor contains a relatively long polyglutamine tract (Giovannucci et al., 1997).
Prostate cancer is the most common cancer in men in many western countries, and the second leading cause of cancer deaths in men. It accounts for more than 40,000 deaths in the US annually. The number of deaths is likely to continue rising over the next 10 to 15 years. In the US, prostate cancer is estimated to cost $1.5 billion per year in direct medical expenses. In addition to the burden of suffering, it is a major public-health issue. Numerous studies have provided evidence for familial clustering of prostate cancer, indicating that family history is a major risk factor for this disease (Cannon et al., 1982; Steinberg et al., 1990; Carter et al, 1993).
Prostate cancer has long been recognized to be, in part, a familial disease with a genetic component (Woolf, 1960a; Cannon et al., 1982; Carter et al., 1992). Numerous investigators have examined the evidence for genetic inheritance and concluded that the data are most consistent with dominant inheritance for a major susceptibility locus or loci. Woolf (1960b), described a relative risk of 3.0 of developing prostate cancer among first-degree relatives of prostate cancer cases in Utah using death certificate data. Relative risks ranging from 3 to 11 for first-degree relatives of prostate cancer cases have been reported (Cannon et al., 1 982; Woolf, 1960b; Fincham et al., 1990; Meikle et al., 1985; Krain, 1974; Morganti et al., 1956; Goldgar et al., 1994). Carter et al. (1992) performed segregation analysis on families ascertained through a single prostate cancer proband. The analysis suggested Mendelian inheritance in a subset of families through autosomal dominant inheritance of a rare (q=0.003), high-risk allele with estimated cumulative risk of prostate cancer for carriers of 88% by age 85. Inherited prostate cancer susceptibility accounted for a significant proportion of early-onset disease, and overall was responsible for 9% of prostate occurrence by age 85. Recent results demonstrate that at least four loci exist which convey susceptibility to prostate cancer as well as other cancers. These loci are HPC1 on chromosome 1q24, (Smith et al., 1996), HPCX on chromosome Xq27-28 (Xu et al., 1998), PCAP at 1q42 (Berthon et al., 1998) and CAPB at 1p36 (Gibbs et al., 1999a). All four suggestions of linkage for prostate cancer predisposition were the result of hints arising from genome-wide searches. Although only the HPC1 linkage has so far been confirmed (Cooney et al., 1997; Neuhausen et al., 1999; Xu and ICPCG, 2000), it is becoming clear that a large number of genes contribute to familial prostate cancer. It also seems clear, both from published hereditary prostate cancer linkage studies and from genotyping of our family resource at the above mentioned loci, that no single predisposition locus mapped to date is by itself responsible for a large portion of familial prostate cancer (Neuhausen et al., 1999; Eeles et al., 1998; Gibbs et al., 1999b; Lange et al., 1999; Berry et al., 2000; Suarez et al., 2000: Goode et al., 2000).
A comparison to the cloning of, and risk profile attributed to, breast cancer susceptibility genes provides an instructive example. The profusion of proposed prostate loci, coupled with minimal confirmation or refined localization following initial publication of these linkages, contrasts sharply with studies of the breast and ovarian cancer susceptibility genes BRCA1 and BRCA2. Linkage to BRCA1 was first published in 1990 (Hall et al., 1990); groups competing to identify this gene moved swiftly from confirmatory studies through efforts to refine the localization to the gene identification in 1994 (Miki et al., 1994). With expanded genomics resources, the time from linkage (Wooster et al., 1994) to complete cloning (Wooster et al., 1995; Tavtigian et al., 1996) of BRCA2 was only slightly more than 1 year. Ongoing mutation screening and careful modeling of age specific and familial risks indicate that these two genes account for virtually all extended breast and ovarian cancer families (Antoniou et al., 2000) and the majority of breast cancer families with more than five cases, especially those that include an early-onset component (Ford et al., 1998).
Even so, a fraction of familial breast cancer risk is manifest in smaller family clusters with average age at diagnosis. While BRCA1 and BRCA2 only account for a portion of this component of breast cancer risk (Peto et al., 1999), there are no published and confirmed linkages based on these types of pedigrees to date. Standard genetic analysis appears to be limited by the problems of low penetrance and genetic complexity. It is possible that analysis of genetic predisposition in families with excess prostate cancer also reflects these issues. As absence of distinction by age at diagnosis/onset would also be consistent with the influence of multiple susceptibility genes harboring only moderate risk sequence variants, one might therefore ask what relative contribution low frequency high risk variants analogous to mutations in BRCA1/2, versus higher frequency, moderate risk sequence variants, make to the population risk of prostate cancer.
Indeed, evidence that moderate risk sequence variants in a number of specific genes contribute to prostate cancer susceptibility is beginning to accumulate. For example, a polymorphic CAG repeat within the androgen receptor open reading frame encoding a variable length polyglutaminc tract shows an inverse relationship between repeat length and the transcriptional transactivation activity of the receptor (Chamberlain et al., 1994; Kazemi-Esfarjani et al., 1995). Accordingly, a series of studies show an association between shorter androgen receptor CAG repeat length and prostate cancer risk (Giovannucci et al., 1997; Stanford et al., 1997), although it is not entirely clear whether the association is with diagnosis of prostate cancer or severity of disease (Bratt et al., 1999). Second, a number of missense variants have been observed in the steroid 5.alpha.-reductase type II gene (SRD5A2), responsible for conversion of testosterone to the more active androgen dihydrotestosterone in the prostate (Makridakis et al., 1997). One of these variants, Ala 49 Thr, has been reported to increase the catalytic activity of the enzyme, and is associated with increased risk of advanced prostate cancer (Makridakis et al., 1999; Jaffe et al., 2000). Finally, several groups have reported an excess of prostate cancer in large BRCA2 pedigrees (Sigurdsson et al., 1997; Breast Cancer Linkage Consortium, 1999), though the relative risk that these mutations confer for prostate cancer is considerably lower than for breast cancer. Further, these effects may be variant specific as association has not been confirmed among men who carry the Ashkenazi BRCA2 founder mutation 6174delT (Wilkens et al., 1999; Nastiuk et al., 1999; Hubert et al., 1999). If these and similar sequence variants play a role in a significant fraction of prostate cancer, then models of the genetic component of familial prostate cancer may need to incorporate both linkage evidence for major susceptibility loci and association evidence for moderate risk sequence variants.
The Utah population provides a unique resource for examining the genetic basis of disease. Extended high risk pedigrees containing many cases can be ascertained as units instead of by expansion from individual probands. While these pedigrees are an extremely powerful resource for linkage studies, they also allow analysis of segregation of moderate risk sequence variants through multiple generations of both cases and their unaffected relatives.
Detection of genetic linkage for prostate cancer susceptibility to a defined segment of a chromosome requires that DNA sequence variants within that chromosomal segment confer the cancer susceptibility. This is usually taken to mean that the causal sequence variant(s) will either alter the expression of one or more linked genes or will alter the function of one of the linked genes. However, detection of the genetic linkage does not necessarily provide evidence for what class of gene (i.e. tumor suppressor, oncogene, or risk modifier) is affected by the causal sequence variant(s).
Most strategies for proceeding from genetic linkage of prostate cancer susceptibility to chromosome 17p to identification of the 17p-linked prostate cancer predisposing gene (HPC2) require precise genetic localization studies to define a discrete segment of the chromosome within which the causal sequence variant(s) must map. Gene identification projects based on precise genetic localization are called positional cloning projects. The general strategy in positional cloning is to find all of the genes located within the genetically defined interval, identify sequence variants in and around those genes, and then determine which of those sequence variants either alter the expression or the function of one (or more) of the associated genes. Segregation of such sequence variants with the disease in the linked kindreds must also be demonstrated. We have executed a positional cloning project in the HPC2 region of chromosome 17p and found a gene, herein named HPC2, germline mutations which predisposes individuals to prostate cancer.